The invention relates to plant regulatory elements that can be used to express various nucleic acid molecules in transgenic plants.
An essential element for genetic engineering of plants is the ability to express genes in various plant tissues. Promoter elements contain specific sequences that guide the binding of proteins necessary for formation of a transcriptional complex, and consequently, allow transcription to begin. Constitutive promoters such as the cauliflower mosaic virus (CaMV) 35S and 19S promoters, as well as Agrobacterium derived promoters such as the nopaline synthase and octopine synthase promoter are widely used for expressing genes in transformed plants.
The invention is based on the identification of regulatory elements of plant fatty acid desaturase genes, which allows any heterologous nucleic acid molecule to be expressed in plant cells and plants. Thus, plants can be produced that have increased nutritional value, a modified fatty acid composition, or a modified carbohydrate, protein, or secondary metabolite content. In addition, the regulatory elements can be used to produce new pharmaceutical products in plants.
In one aspect, the invention features an isolated nucleic acid molecule including a regulatory element of a fatty acid desaturase gene, wherein the regulatory element is located 5xe2x80x2 of the fatty acid desaturase gene in a naturally occurring genome. The fatty acid desaturase gene can be, for example, a Brassica fad2D gene or a Brassica fad2F gene.
The invention also features an isolated nucleic acid molecule having the nucleotide sequences of SEQ ID NO:4, SEQ ID NO:5, nucleotides 5113 to 5796 of SEQ ID NO:4, or nucleotides 3197 to 3867 of SEQ ID NO:5, as well as the complementary sequence of such nucleic acid molecules. The nucleic acid molecule also can be at least 100 nucleotides in length (e.g., 400, 600, or 800 nucleotides) and have at least 70% sequence identity (e.g., 80%, 90%, or 95%) to such nucleic acid molecules. In some embodiments, the isolated nucleic acid molecule includes nucleotides 5113 to 5796 of SEQ ID NO:4 or nucleotides 3197 to 3867 of SEQ ID NO:5.
The invention also features a nucleic acid construct. The construct includes a first nucleic acid molecule operably linked to a second nucleic acid molecule heterologous to the first nucleic acid molecule. The first nucleic acid molecule includes a regulatory element of a fatty acid desaturase gene, wherein the regulatory element is located 5xe2x80x2 of the fatty acid desaturase gene in a naturally occurring genome. The first nucleic acid molecule promotes expression of the heterologous second nucleic acid molecule, which can encode, for example, a ribozyme or a polypeptide, such as a polypeptide that confers herbicide resistance.
The nucleic acid construct further can include a third nucleic acid molecule operably linked to the 3xe2x80x2 end of the second nucleic acid molecule. The third nucleic acid molecule includes a 3xe2x80x2 untranslated region of a fatty acid desaturase gene, wherein the 3xe2x80x2 untranslated region is located 3xe2x80x2 of the fatty acid desaturase gene in a naturally occurring genome. The third nucleic acid molecule can include a nucleic acid molecule having the nucleotide sequence of SEQ ID NO:6, SEQ ID NO:7, nucleotides 1-894 of SEQ ID NO:6, or nucleotides 1-817 of SEQ ID NO:7, as well as the complementary sequence of such molecules. The nucleic acid molecule can be at least 100 nucleotides in length and have at least 70% sequence identity to a nucleic acid molecule having the nucleotide sequence of nucleotides 1-894 of SEQ ID NO:6 or nucleotides 1-817 of SEQ ID NO:7.
The nucleic acid construct further can include a fourth nucleic acid molecule operably linking the first and second nucleic acid molecules, wherein the fourth nucleic acid molecule is a transit peptide or an intron.
In another aspect, the invention features an isolated nucleic acid construct including a first nucleic acid molecule operably linked to a second nucleic acid molecule heterologous to the first nucleic acid molecule. The first nucleic acid molecule comprises a3xe2x80x2 untranslated region of a fatty acid desaturase gene, wherein the3xe2x80x2 untranslated region is located3xe2x80x2 of the fatty acid desaturase gene in a naturally occurring genome.
The invention also features transformed plant cells and transgenic plants. The transformed plant cells and transgenic plants include a nucleic acid construct. The nucleic acid construct includes a regulatory element of a fatty acid desaturase gene operably linked to a heterologous nucleic acid, wherein the regulatory element promotes expression of the heterologous nucleic acid, and wherein the regulatory element is located 5xe2x80x2 of the fatty acid desaturase gene in a naturally occurring genome. The transgenic plant can be a dicot (e.g., alfalfa, soybean, rapeseed, or sunflower) or a monocot (e.g., corn, wheat, rye, rice, or sorghum). Seeds from the transgenic plant also are featured.
In another aspect, the invention features plasmid pMB102, represented by ATCC accession number PTA-2536, and plasmid pMB103, represented by ATCC accession number PTA-2535.
Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although methods and materials similar or equivalent to those described herein can be used to practice the invention, suitable methods and materials are described below. All publications, patent applications, patents and other references mentioned herein are incorporated by reference in their entirety. In case of conflict, the present specification, including definitions, will control. In addition, the materials, methods, and examples are illustrative only and not intended to be limiting.
Other features and advantages of the invention will be apparent from the following detailed description, and from the claims.